All photosynthetic eukaryotic cells contain chloroplasts that use the radiant energy of sunlight to convert carbon dioxide and water into carbohydrates. As a byproduct of photosynthesis, oxygen gas is also released into the atmosphere through tiny openings in the leaves called stomata. The carbohydrates produced by photosynthesis provide us with an important energy source, while the oxygen is a critical component of the air we breathe.
The reactants and products of photosynthesis have been known since the early 1800s. At first, it was generally assumed that the oxygen released as a byproduct of photosynthesis came from the carbon dioxide. However, the question was not definitely answered until the early 1940s, and the result was surprising to many.
In plants, water enters through the roots while atmospheric carbon dioxide enters through openings in the leaves called stomata. Oxygen and water are released by the plant through these same openings. Photosynthesis takes place within the leaves, in specialized organelles called chloroplasts.
The basic process of photosynthesis can be summarized using the equation shown here: carbon dioxide, water, and light energy combine to yield the products of sugar, oxygen, and water.
Of course, knowing the reactants and products does not tell us the specific nature of the photosynthetic reactions that take place. For example, where does the oxygen that is released into the atmosphere by plants come from? Does it come from the carbon dioxide, or from the water?
This question was answered in the 1940s by Samuel Ruben and colleagues at UC Berkeley using isotopes of oxygen. The common isotope of oxygen is oxygen-16. By using the oxygen-18 "heavy" isotope of oxygen to label either the carbon dioxide or water reactants, the source of the oxygen released into the atmosphere can be determined.
Let's begin by labeling the carbon dioxide reactant with the oxygen-18 isotope. Our data will consist of the 18O/16O isotope ratio measured in the evolved oxygen as compared to the starting ratio in the CO2 reactant (and unlabeled H2O). Note that not all of the CO2 will contain the heavy isotope.
Now let's repeat the experiment by labeling the H2O reactant. Again, our data will consist of the 18O/16O isotope ratio measured in the evolved oxygen as compared to the starting ratio in the H2O reactant.
What can we conclude from this experiment? Is the source of oxygen released by the plants derived from the carbon dioxide or water reactants?
When CO2 was labeled with 18O, none of the oxygen released was labeled. However, when 18O-labeled water was used as a reactant, the oxygen released was labeled with the heavy isotope. Therefore the source of the oxygen in the O2 released by the plant is water.
One of the first uses of an isotopic tracer in biology was in the determination of the source of the oxygen produced by photosynthesis. Using a heavy isotope of oxygen, 18O, to label one of the two reactants in the photosynthetic process, Samuel Ruben and collegues at UC Berkeley were able to determine that the oxygen gas came not from carbon dioxide, but from water.